KANPHOS: Newly developed database for kinase-associated protein phosphorylation eases neural signaling research

Signal transduction, or cellular communication involving chemical and physical signals transmitted by signal molecules in the extracellular and intracellular environments, leads to the generation of a signal cascade, and is essential for correct execution of biological processes. This process is facilitated by the mechanism of phosphorylation and dephosphorylation—addition or removal of phosphate groups from specific substrate molecules by a family of enzymes called kinases. Hundreds of such enzymes have been identified in humans, which play an important role in the maintenance of a variety of key functional processes. This is especially true for proper neurological functioning, with deficiencies in signaling pathways related to a variety of neurological disorders.

While understanding the components and molecular mechanisms behind these signaling strategies is important, the exact involvement of individual kinases and substrates has not been clearly elucidated. In order to bridge this gap, scientists from Fujita Health University, Toyoake, and their multi-institutional colleagues designed a database called Kinase-Associated Neural PHOspho-Signaling (KANPHOS), which provides extensive data on different aspects of phosphorylation in the brain, such as signaling strategies, phosphoproteins involved, phosphorylation sites, and participant kinases. Prof. Kozo Kaibuchi, lead researcher on the study published in Cells, explains the team’s motivation behind conducting this study, “Protein phosphorylation is crucial for revealing how organs and cells function and what happens during some diseases. Accumulating studies have identified various signaling pathways but there are thousands of proteins that can be phosphorylated, suggesting that there are possibly countless unknown signaling pathways that involve protein phosphorylation.”

This newly developed database contains information about new phosphoproteins, phosphorylation sites of interest, as well as molecular species specificity. The database is built on the software platform XOOPS module for Neuroinformatics (XooNIps), which allows for constructing a web-based database that can handle a large set of metadata in different formats. Moreover, this can be accessed worldwide, which increases the range of its possible impact.

Constructing such an extensive database involved collecting information from different sources. The team manually curated existing datasets to supplement the database with information on already available phosphoproteomic data. Additionally, the researchers employed advanced techniques to further determine novel kinase-oriented protein phosphorylation information, such as the in vitro kinase interacting substrate screening method, which involved determining protein substrates through a selection process involving interactions with the catalytic (enzymatically active) domain of specific kinases, followed by their characterization with liquid chromatography/mass spectrometry (LC-MS). An in vivo technique called the kinase-oriented substrate screening method involved identifying substrates by treating tissue samples with agonists (activators) and kinase inhibitors, followed by enrichment and characterization of the identified phosphoproteins via LC-MS.

The team’s focus while designing the database was on user friendliness, besides accuracy. This was reiterated by Prof. Kaibuchi, who, speaking on behalf of co-authors Taku Nagai and Junichiro Yoshimoto from Fujita Health University, Mutsuki Amano from Nagoya University, and Takayuki Kannon from Kanazawa University, mentions, “KANPHOS is designed so that users can easily access information on genes, signaling pathways, physiological functions, and diseases associated with protein phosphorylation. This functionality will promote data-driven studies on molecular mechanisms in the brain.” Thus, the database allows for searches based on phosphorylated proteins, specific kinases, or the pathways themselves. There are also options for advanced searches, which could include searches based on gene/protein characteristics, pathways, experimental conditions, and other specific requirements. Another useful feature added is linking information from other databases directly into the webpage, ensuring a complete representation of information on a specific protein.

This database has immense potential to facilitate research in elucidating signaling pathways. The team was able to demonstrate its efficacy by exploring the different components of the adenosine A2A receptor signaling pathway mediated by adenosine—an endogenous purine nucleoside that modulates neural function—in murine brain. The database was instrumental in elucidating the specifics of the MEK-MAPK pathway driven by adenosine, and also helped in identifying a vital substrate in this process, Arhgap21. Prof. Kaibuchi is optimistic about the benefits of this database, claiming that “The information infrastructure provided by the database may help elucidate the patho-etiology and pathophysiology of neuropsychiatric disorders.”

This tool establishes the usefulness of novel information technologies in data-driven research, especially in the neurosciences, with great scope for developing therapeutic strategies for debilitating neurological disorders in future.

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Reference

Title of original paper: KANPHOS: A Database of Kinase-Associated Neural Protein Phosphorylation in the Brain

Journal: Cells

DOI: https://doi.org/10.3390/cells11010047

About Fujita Health University

Fujita Health University is a private university situated in Toyoake, Aichi, Japan. It was founded in 1964 and houses one of the largest teaching university hospitals in Japan in terms of the number of beds. With over 900 faculty members, the university is committed to providing various academic opportunities to students internationally. Fujita Health University has been ranked eighth among all universities and second among all private universities in Japan in the 2020 Times Higher Education (THE) World University Rankings. THE University Impact Rankings 2019 visualized university initiatives for sustainable development goals (SDGs). For the “good health and well-being” SDG, Fujita Health University was ranked second among all universities and number one among private universities in Japan. The university will also be the first Japanese university to host the "THE Asia Universities Summit" in June 2021. The university’s founding philosophy is “Our creativity for the people (DOKUSOU-ICHIRI),” which reflects the belief that, as with the university’s alumni and alumnae, current students also unlock their future by leveraging their creativity.

Website: https://www.fujita-hu.ac.jp/en/index.html

About Professor Kozo Kaibuchi from Fujita Health University

Prof. Kozo Kaibuchi graduated from Kobe University, School of Medicine to become a medical doctor in 1980 and obtained a PhD in Biochemistry from the same University in 1984. He is currently the Director of the International Center for Brain Science, Fujita Health University. His interests lie in the study of molecular pathogenesis in psychiatric disorders like schizophrenia. He has been at the forefront for the development of a novel phospho-proteomic approach to comprehensively screen for substrates of protein kinases including Rho-kinase, protein kinase A, and MAP-kinase. He is also working on approaches that analyze how dopamine, adenosine, and acetylcholine regulate neuronal functioning.